Imaging of the mounted larvae was performed using a custom-made upright fluorescence microscope. We imaged either the entire zebrafish larva (using EC Plan-NEOFLUAR 2.5x/0.085 M27, Carl Zeiss Microscopy) or only the encephalic region (with EC Plan-NEOFLUAR 5x/0.16 M27, Carl Zeiss Microscopy). Fluorescence was excited in wide-field configuration using an LED with emission centred at 470 nm (M470L3, Thorlabs), followed by an excitation band-pass filter (469/35 nm, FF01–469/35, Semrock). The epifluorescence detection was implemented with a dichroic mirror (DC FF495-DI02, Semrock) and an emission band-pass filter (525/39 nm, MF525-39, Thorlabs). The fluorescence signal was recorded with a sCMOS camera (OrcaFLASH 4.0, Hamamatsu Photonics) with 16-bit dynamic range.
To perform high-throughput measurements the excitation LED was equipped with a diffuser to produce an even illumination over an area of about 50 cm2, covering all the multi-well plate. In order to image such a large area, a wide-angle lens (MVL8M23, Thorlabs) with 8 mm focal length was mounted directly on the camera.
To perform high-throughput measurements the excitation LED was equipped with a diffuser to produce an even illumination over an area of about 50 cm2, covering all the multi-well plate. In order to image such a large area, a wide-angle lens (MVL8M23, Thorlabs) with 8 mm focal length was mounted directly on the camera.